Genomic instability of osteosarcoma cell lines in culture: impact on the prediction of metastasis relevant genes.

Institute of Molecular Life Sciences, University of Zurich, Zurich, Switzerland.

Abstract

BACKGROUND:

Osteosarcoma is a rare but highly malignant cancer of the bone. As a consequence, the number of established cell lines used for experimental in vitro and in vivo osteosarcoma research is limited and the value of these cell lines relies on their stability during culture. Here we investigated the stability in gene expression by microarray analysis and array genomic hybridization of three low metastatic cell lines and derivatives thereof with increased metastatic potential using cells of different passages.

PRINCIPAL FINDINGS:

The osteosarcoma cell lines showed altered gene expression during in vitro culture, and it was more pronounced in two metastatic cell lines compared to the respective parental cells. Chromosomal instability contributed in part to the altered gene expression in SAOS and LM5 cells with low and high metastatic potential. To identify metastasis-relevant genes in a background of passage-dependent altered gene expression, genes involved in "Pathways in cancer" that were consistently regulated under all passage comparisons were evaluated. Genes belonging to "Hedgehog signaling pathway" and "Wnt signaling pathway" were significantly up-regulated, and IHH, WNT10B and TCF7 were found up-regulated in all three metastatic compared to the parental cell lines.

CONCLUSIONS:

Considerable instability during culture in terms of gene expression and chromosomal aberrations was observed in osteosarcoma cell lines. The use of cells from different passages and a search for genes consistently regulated in early and late passages allows the analysis of metastasis-relevant genes despite the observed instability in gene expression in osteosarcoma cell lines during culture.

The number of differentially expressed genes at different significance levels was normalized to calculated doublings in order to compare the three cell line systems. Note the different y-axis scales for each cell line system. fdr; false discovery rate.

(A) CN gains (yellow) and losses (blue) compared to normal diploid human genome were analyzed in SAOS and LM5 cells of early and late passages and compared to published data of SAOS cells (GSM170249; []) using arrayMap as described in Methods. (B) Statistics of CN gains and losses compared to normal human diploid genome. (C) Total CN differences between early and late passages of SAOS and LM5 cells.

Indicated genes are up-regulated (>2-fold; p<0.05) in at least three passage comparisons. Genes up-regulated in four passage comparisons are shown in bold. Genes marked in red are up-regulated in all three metastatic cell lines. Genes marked in purple, blue and green are shared by LM5 and LM8, LM5 and143B and LM8 and 143B, respectively.